Key words: GTPases; oncogenes; transformation; transcription factors; signal transduction; apoptosis; metastasis; carcinogenesis

Summary
Ras and Rho GTPases are among the best studied signaling molecules in molecular biology. Essential cellular processes such as, cell growth, lipid metabolism, cytoarchitecture, membrane trafficking, transcriptional regulation, apoptosis and response to genotoxic agents, are directly modulated by different members of this superfamily of proteins. Not until recently, we have started to understand the physiological implications of Ras and Rho GTPases, linking them to processes such as embryonic development, tissue remodeling, tumorigenesis and metastasis. In this sense, uncontrolled activation, due to overexpression of different members of the Rho family in a variety of tissues, leads to uncontrolled proliferation and invasiveness of human tumors. In this review, an attempt to briefly integrate recent findings in transcriptional regulation by Rho GTPases in the context of carcinogenesis and metastasis as well as apoptosis is made.

1. Introduction
Over 100 members of the Ras superfamily of GTPases have been found so far. Based on both sequence homology and function, they have been subdivided into at least six families: Ras, Rho, Rab, Arf, Ran and Rad/Gem [1-3]. Each family, in turn, is comprised of several members with distinct expression, cellular localization and biological activities. The Rho family includes RhoA, RhoB, RhoC, Cdc42, TC10, Rac1, Rac2, Rac3, RhoG, Rho6/Rnd1, Rho7/Rnd2, Rho8/Rnd3, RhoE, RhoD, RhoH and TCF. These proteins cycle between an active GTP-bound state and an inactive GDP-bound state in a tightly regulated manner controlled by several regulatory proteins: guanine exchange factors (GEFs); GTPase-activating proteins (GAPs); and Guanine Nucleotide Dissociation Inhibitors (GDIs).

Both Ras and Rho GTPases mediate key cellular processes in response to diverse stimuli, such as cell growth, apoptosis, lipid metabolism, cytoarchitecture, membrane trafficking, and transcriptional regulation. However, the negative aspect of these multifunctional proteins arises in the context of scenarios that cause their constitutive activation (i.e. point mutations or overexpression) and render them insensitive to regulatory signals. In this case, these GTPases trigger specific signals that lead to uncontrolled cell growth, enhanced angiogenesis, inhibition of apoptosis and genetic instability, all which result in tumor development.

The roles of Ras and Rho GTPases in cell cycle, differentiation, and regulation of cytoskeleton have been extensively reviewed recently [1-5]. Both the upstream stimuli that activate Rho members and the time frame of activation of these proteins during development are starting to be unraveled [2]. As well, a whole new set of effectors to these proteins have been cloned, and their specific roles are being studied both in the context of each GTPase alone and at the level of functional interaction among different members of this family of GTPases [4]. Despite the amount of evidence that points out a role of Rho GTPases in transformation and metastasis, as well as regulation of apoptosis, these effects have been commonly underscored. Moreover, difficulties in unraveling Rho functions may be due to their complexity in switching a large variety of signaling cascades both at the membrane/cytosolic and nuclear level. Understanding the integration of signals at the nuclear level may provide the clue for proper interpretation of their evasive biological functions. In this review, a brief overlook of transformation and apoptosis induced by different members of Rho GTPases and their relationship with transcriptional regulation is attempted. Due to space limitations, we will mostly focus on the archetypes RhoA, Rac1 and Cdc42.

2. Rho GTPases and regulation of transcription factors
Rho GTPases trigger different intracellular pathways that lead to the activation of several transcription factors (TFs) and nuclear signaling (Fig. 1). However, little is known yet on the intricate differences among all Rho proteins involved, the precise optimal conditions for the activation of specific TFs, nor their specific targets. All three, RhoA, Rac1, and Cdc42 activate SRF and NF-kB, as well as TFs dependent on the activity of JNK1 and p38. Such substrates to these kinases include ATF-2, ELK [6-7], PEA3 [8-10], Max and CHOP/GADD153 [11-12].

An example of the complexity of the activation of TFs by Rho proteins is the stimulation of the transcription of c-fos proto-oncogene [13-14]. This regulation is dependent on the SRE element present at the proximal promoter region of the gene, and independent of TCF (ternary complex factor) [14]. Activation of the SRE element of the c-fos promoter by RhoA, but not Rac1 and Cdc42Hs, involves the formation of a multiprotein transcriptional complex comprised of SRF, NF-kB and C/EBPb [15]. Thus, activation of NF-kB by RhoA does not exclusively promote its nuclear translocation and binding to the specific kB sequences, but also involves the regulation of the transcriptional activity of the c-fos Serum Response Factor (SRF). In fact RelA and p50 NF-kB subunits cooperate with the transcription factor C/EBPb in the transactivation of the SRE reporter. RhoA increases the levels of C/EBPb protein, facilitating the functional cooperation between NF-kB, C/EBPb and SRF proteins. Therefore, whereas modulation of NF-kB occurs via IKBa, the regulation of C/EBPb appears to be by incrementing its proteins levels. These results strengthen the pivotal importance of the Rho family of small GTPases in signal transduction pathways which modulate gene expression and reveal that NF-kB and C/EBPb transcription factors are accessory proteins for the RhoA-linked regulation of the activity of the serum response factor (SRF).

Finally, other authors have suggested that transcriptional activation of SRF is susceptible to actin rearrangements through the Rho effector LIM Kinase [16]. SRF could be linking actin reorganization to RhoGTPases since it transcriptionally regulates several cytoskeletal-related genes such as vinculin, cytoskeletal actin, and SRF itself. However, the physiological effect of this SRF-dependent effect in the context of RhoGTPases remains to be determined. ,p> Besides the SRE element, the promoter region of c-fos contains several other DNA elements specific for other TFs such as AP1, TCF, CREB and Stat [17]. The role, if any, of these nuclear proteins in Rho-mediated induction of c-Fos is mostly unknown. Two recent findings suggest that these sites may be relevant to Rho function. First, transcription of the Stat-inducible element (SIE) of c-fos is highly induced by RhoA, and to a less extent by Cdc42Hs, but not Rac1, in human embryonic kidney cells (293T), Chinese Hamster Ovary cells (CHO) and Buffalo Rat Kidney cells (BRL) [S. Aznar et al, submitted]. The SIE element lies adjacent to the SRE, and is responsive to homo- or heterodimers of Stat1 and Stat3 [18]. In fact, SIE activation by RhoA is dependent on Stat3, and upon oncogenic Rho-signalling Stat3 is both tyrosine and serine phosphorylated, hence transcriptionally active. Accordingly, it has been described that Rac1 binds to Stat-3 through its effector domain, and this interaction triggers tyrosine phosphorylation and activation of Stat3 in COS-1 and Rat cells by a JAK2-dependent mechanism [19]. Whereas tyrosine phosphorylation of Stat3 upon RhoA and Rac1 activation is via a JAK2 pathway, in the context of RhoA serine phosphorylation of Stat3 is carried by JNK, but not ERK1 or p38 [S. Aznar et al, submitted]. Thus, modulation of Stat3 by Rho GTPases might be a common event, which displays tissue specificity.

3. Rho GTPases: a role in transformation and metastasis
The relationship of Rho proteins with cell transformation and human cancer is building up strongly. Although it was shown very early that Rho GTPases have transforming properties both in vivo and in vitro [20-21], this finding was shadowed by the discovery of their role in regulating the cytoskeleton. Recent studies have reinforced the oncogenic potential of Rho proteins [4, 22-25]. In addition, several studies suggest that Rho GTPases might be commonly overfunctional in human cancers [25-28]. Finally, expression of activated Rho proteins is sufficient for the acquisition of the full metastatic phenotype [29-30]. To further strengthen the relevance of Rho proteins in the carcinogenic process, it has been shown that Ras-dependent transformation requires functional Rho proteins, including RhoA, Rac1 and Cdc42Hs [31-33]. Thus, Ras and Rho GTPases-mediated transformation can no longer be seen as separate events, and the pathways elicited by both families form a complex weave that finally results in tumorigenesis and metastasis.

An increasing interest in the mechanisms whereby these proteins trigger the intracellular cascades that lead to transformation has been observed in the past decade. In this sense many effectors to Rho GTPases are involved in transformation and metastasis [4, 30] (Fig. 2). Furthermore, in some cases the modulation of transcription has been directly linked to transformation (Fig. 2). In this sense, different studies have suggested a role for PAK, an effector to Rac and Cdc42 in transformation downstream of these GTPases. For instance, several groups have reported a role for PAK in Rac and Ras-mediated signalling and transformation [34-35]. Also, a mutant p65PAK that lacks the kinase domain but retains the ability to bind to Rac or Cdc42 inhibits Ras-dependent transformation of rat 3Y1 and Rat-1 cells [34, 36]. As well, combinations of Rac/Raf, Ras/Raf, and Rho/Raf, show synergism in both ERK activation and transformation in a PAK-dependent manner.

However, there is some controversy around PAK-dependent transformation, since Rac-induced transformation of NIH3T3 fibroblasts is independent of PAK1 [37]. However, a possible reason is that p65PAK might be necessary for transformation in the context of Ras-Rac pathway, but not for Rac-mediated transformation independently of Ras. Consistent with this hypothesis is the fact that a dominant active mutant of PAK itself does not elicit Schwann cell transformation, but a dominant negative mutant of PAK can inhibit transformation induced by Ras in these neuronal cells [38]. On the other hand, hyperactive Rac3 and PAK are necessary for human breast cancer cell DNA synthesis, and tumor growth in a JNK-independent manner [35].

Rho GTPases stimulate both p38 and JNK activity [39]. Although PAKs have been implicated in JNK- and p38-activation downstream of Rac and Cdc42 and constitutively active mutants of PAK can stimulate both JNK and p38 activities [40-42], mutants of Rac that fail to bind PAK remain capable of inducing JNK activity [37]. In any case, both Rac and Cdc42 can use different effectors to activate the JNK/p38 MAP kinases pathways. One candidate is the Rac effector POSH (Plenty Of SH3 domains) [43] which when expressed in COS-1 cells mediates Rac1-induced JNK activation and NF-kB translocation to the nucleus. As well, two proteins termed MEKK4 and MLK3 are activated by both Rac and Cdc42 and ultimately lead to JNK and p38 activation [44-47]. Interestingly, MLK3 links Rho GTPases to NF-kB activation [48], and is involved in Rac/Cdc42-mediated transformation of NIH3T3 cells in a MEK-dependent fashion [49].

In addition to their role in transformation, MLK3 and MEKK4-dependent signals might lead to invasiveness and metastasis of tumor cells. This is predicted from the fact that activation of endogenous p38 is necessary for the metastatic phenotype of breast carcinoma cells, by promoting transcription and mRNA stabilization of the urokinase plasminogen activator (uPAR) gene [50-51]. Interestingly, a pyrimidazole derivative, SB203580, which specifically inhibits p38, abrogates the metastatic capacity of these cells [52]. However, its role in Rac1/Cdc42 induced metastasis is unclear since neither induce transcription of the uPAR gene [52]. On the other hand, a role for Ras and RhoA in uPAR regulation has been described [52]. Both these proteins in their oncogenic versions stimulate transcription of the uPAR gene promoter region, leading to increased levels of its protein product. This seems to be regulated by extracellular matrix signals (ECM) such as laminin or fibronectin [53], which lead to increased metalloproteases (MMP) and uPAR that results in enhanced motility and invasiveness of cells.

Interestingly, the proximal region of the urokinase plasminogen receptor contains an AP-1 site and a kappaB site specific for NF-kB [54-55]. Hence, Rho GTPases might enable enhanced invasiveness and metastasis of tumor cells via an MMP- uPAR pathway at the transcriptional level via JNK and NFkB.

All three Rho GTPases prototypes, RhoA, Rac1 and Cdc42, as well as Ha-Ras efficiently induce nuclear translocation and activation of NFkB [15, 56-57]. Consistent with a role of transcription in Rho-mediated transformation, this transcription factor seems to be transforming in the context of these proteins [58-62]. Accordingly, NFkB activation is necessary for Dbl and Dbs-induced transformation via Rho GTPases [62].

One effector to RhoA, Rac1, and Cdc42 involved in uPAR regulation and transformation is Phospholipase D (PLD). For instance, PLD activation in v-Raf transformed cells is inhibited by dominant negative mutants of both Ral and Rho, suggesting a role for the latter in Ras-mediated activation of PLD1 [63]. Whereas, PLD contributes to transformation by promoting a signal at the transcriptional level is currently unknown.

Another effector to Rho that has been tightly linked to transformation and metastasis is ROCK [64]. In this sense, a ROCK specific inhibitor, Y-27632, is capable of inhibiting growth of Ras and Rho-dependent tumors in vivo [65-66]. Interestingly, this drug is not capable of inhibiting Rho-mediated serum response factor (SRF) activation, and transcription of c-fos promoter, suggesting that these Rho-induced effects might not be indispensable for transformation [64, 66].

In addition, Y-27632 also inhibits transcellular invasion of rat MM1 hepatoma-induced tumors in vivo [66], and a dominant negative mutant of ROCK substantially abrogates the invasive phenotype of these cells. Interestingly, Y-27632 treatment of rat MM1 hapatoma cells results in inhibition of Rho-mediated actomyosin cytoskeletal changes, hallmarks of Rho activity suggesting that Rho-mediated transformation is independent on cytoskeletal changes, whereas the invasive phenotype elicited by Rho is dependent on actin reorganization.

As mentioned above, Stat3 lies downstream of both Rho and Rac in different cellular lines. This pathway might be relevant to Rho-mediated transformation since both oncogenic RhoA (Q63L) and Stat3 synergizes in anchorage-independent growth stimulation and a dominant negative mutant of Stat3 abolishes anchorage-independent growth elicited by RhoA (QL) transfectants [Aznar et al, unpublished data]. Interestingly, another small GTPase, RalA in the context of EGF elicits a signaling pathway similar to that induced by RhoA, since it activates Src, which leads to activation of Stat3 [67]. However, whether Stat3 is physiologically relevant in human tumors that contain elevated levels of RhoA remains to be elucidated. In addition, the specific effects carried out by Stat3 in the context of RhoA, as well as its relationship with other Stat proteins and/or transcription factors such as NFkB or AP-1 known to be involved in Rho-induced oncogenesis needs to be determined.

Finally, RhoA, Rac1 and Cdc42, activate the serum response factor (SRF) [68]. Whereas both Rac and Rho induce SRE chromosomal templates to some extent [69], oncogenic Cdc42 seems to most efficiently stimulate transcription of these [70]. Nevertheless, there is a great body of evidence that suggests that this transcription factor might not be involved in Rho-mediated transformation. In this sense, whereas ROCK, Dbl or RhoB can activate SRF, this activation is independent of their transforming activity [64, 71]. However, mutants of Rho in its effector binding domains have resulted in contradictory results [72-73].

4. Regulation of programmed cell death by Rho GTPases
The first clue for a role of Rho GTPases in apoptosis came from in vivo tumorigenic studies in nude mice injected with fibroblasts transformed with Rho genes [30]. Tumors generated by stable, Rho-transformed cell lines displayed a high apoptotic index, suggesting that Rho GTPases could be triggering signals that ultimately lead to apoptosis. Accordingly, it was later reported that overexpression of the human genes rho A, rho C and rac 1 induce apoptosis upon serum deprivation in NIH 3T3 fibroblasts and human erythroleukemia K562 cells [74]. In addition, two GEFs for Rho proteins with oncogenic properties, vav and ost, trigger apoptosis under similar conditions [75-76]. In keeping with these observations, transgenic mice carrying an oncogenic version of Rac2 show increased apoptosis in the thymus, which constitutes the first in vivo role of a Rho GTPase in apoptosis [77].

In an effort to investigate the specific factors that induce Rho-mediated apoptosis and its downstream cellular pathways that ultimately lead to the specific changes that take place during cell death, others and we have identified several key players in this process. For instance, a dominant negative mutant of Rac1 (N17) can efficiently abrogate TNFa-induced apoptosis in U937 cells [77]. Furthermore, under serum-starved conditions Rac1 induces complementary signals in NIH3T3 fibroblasts that result in an increase of ceramide levels and Fas ligand (FasL) gene expression that ultimately lead to apoptosis [78]. Whereas, regulation of FasL occurs at the transcriptional level involving both JNK as well as NF-kB-dependent signals, the mechanism responsible for production of ceramides is still unknown. However, both ceramides and FasL production are necessary for apoptosis to take place.

In keeping with these observations, both protein synthesis and caspase-3 activity are required for this process. Surprisingly, no release of cytochrome C from mitochondria, but a rapid increase in mitochondrial membrane potential and the production of reactive oxygen species (ROS) take place, which correlate with the ability of Rac1 to induce apoptosis. Thus, Rac-induced apoptosis takes place by a complex mechanism involving the concurrent generation of ceramides and the de novo synthesis of FasL.

Again, modification in transcriptional regulatory elements by Rac1 plays a critical role in its biological function. However, as discussed above, both JNK and NF-kB are implicated in transformation downstream of Rho GTPases, suggesting that both these signals might tilt the balance towards apoptosis or transformation depending on the extracellular signals (i.e. serum deprivation versus serum or growth factors).,p> Indirect evidence is also building up to support a role of Rho proteins in apoptosis under specific conditions. PAK2 is a substrate to caspases and might be responsible for some of the morphological and structural changes that occur during apoptosis [79-80]. Similarly, a RhoA effector PKN is proteolytically cleaved and activated by caspase-3 upon Fas ligand, staurosporin or etoposide treatment of Jurkat and U937 cells [81]. As well, PRK2 cleaved in early stages of apoptosis, binds to and prevents phosphorylation of Akt in vivo at serine 473 and threonine 308 [82]. Since, phosphorylation of Akt is necessary for its full activation; PRK2 inhibits Akt downstream signaling and abrogates its anti-apoptotic effects. At last, overexpression of POSH in NIH3T3 cells induces apoptosis [43]. Less is known about the specific signals induced during apoptosis by other GTPases. In a manner similar to Rac1, overexpression of RhoA in different cell lines induces an increment of ceramide levels necessary for Rho-induced cell death [77]. Furthermore, this effect is independent on p53 activity but dependent on modulation of anti-apoptotic Bcl-2. However, whether Rho regulates transcription of FasL has not been determined yet. In addition, Cdc42 has been shown to induce apoptosis in Jurkat cells, through a protein kinase cascade leading to stimulation of c-Jun amino terminal kinase (JNK) [83]. Interestingly, stress response- and ceramide-induced apoptosis involves a Ras/ceramide-activated protein kinase as well as a Rac, Raf-1 and MEK pathways [84]. In this pathway, Akt (PKB) is inactivated causing the phosphorylation and activation of pro-apoptotic Bad, which executes its function by binding its partner Bcl-2. Interestingly, this results in the release of cytochrome c from mitochondria to the cytosol triggering cell death [84]. Hence, different GTPases might trigger different apoptotic pathways under specific signals.

Given how recent these observation are, no aims have been made yet to study the physiological role(s) of apoptosis induced by small GTPases in different scenarios, such as development, or different pathologies like cancer. The role of some effectors to Rho GTPases in apoptosis such as PKN, PAK or POSH has been discussed above. Whether these effectors downstream of Rho mediate apoptosis in natural occurring human tumors is unknown. This is of great interest since these proteins could be important players in anoikis, a necessary condition for tumor cells to metastasize. In such case, an intensive study of the microenvironment of the tumoral cells within the tumor (i.e. extracellular matrix signals, autocrine secretions, hypoxia or angiogenesis) is necessary to determine which factors alter the equilibrium between increased transformation or apoptosis induced by Rho GTPases.

5. Conclusions
Our understanding of the roles that Rho proteins may play in transformation and apoptosis, although incipient are now being strongly established. The involvement of signaling cascades where transcription factors are key players is evident. These findings predict a new area of intense research for the old, but always renovated family of Rho proteins. Much more attention will be paid to their participation in signaling cascades leading to nuclear events. The biological read out for these proteins will be focused on their effects on cell growth, and their participation in transformation and metastasis. Finally, a role in apoptosis for Rho proteins opens another battlefield for new insights in development in addition to carcinogenesis.

Acknowledgments
This work was supported by the following Grants: Grant 2FD97-0647 from CICYT, Grant 99/0817 from FIS, Grant 2FD97-1569 from CICYT, Grant 08.1/0045.1/98 from Consejería de Educación of Comunidad de Madrid, and Grant 0112.C02.01 from CICYT.

References
1. I. M. Zohn, S. L. Campbell, R. Khosravi-Far, K. L. Rossman, C. J. Der, Rho family proteins and Ras transformation: the RHOad less traveled gets congested. Oncogene 17(11 Reviews), 1998, 1415-38
2. L. Van Aelst and Crislyn D´Souza-Schorey, RhoGTPases and signaling networks.Genes and Dev 11, 1997, 2295-2322
3. M. Malumbres, A, Pellicer, Ras signaling in cell cycle regulation in its role in tumor development. Rev. Oncología 1, 1999, 66-76
4. S. Aznar and J. C. Lacal, Searching New Targets for Anticancer Drug Design: The families of Ras and Rho GTPases and their effectors. Prog Nucl Acid Res and Mol Biol. 2001, In press
5. D. Bar-Sagi, A. Hall, Ras and Rho GTPases: a family reunion. Cell 13, 2000, 227-38
6. H. Gille, T. Strahl, P. E. Shaw, Activation of ternary complex factor Elk-1 by sterss-activated protein kinases. Curr Biol 5, 1995, 1191-1200
7. S. Gupta, D. Campbell, B. Derijard, R. J. Davis, Transcription factor ATF-2 regulation by the JNK signal transduction pathway. Science 267, 1995, 389-393
8. B. J. Pulverer, J. M. Kyriakis, J. Avruch, E. Nikolakaki, J. R. Woodget, Phosphorylation of c-jun mediated by MAP kinases. Nature 353, 1991, 670-674
9. B. Derijard, J. Raingeaud, T. Barrett, I. H. Wu, J. Han, R. J. Ulevitch, R. J. Davis, JNK1: A protein kinase stimulated by UV light and Ha-Ras that binds and phosphorylates the c-jun activation domain. Cell 76, 1994, 1025-1037
10. R. C. O´Hagan, R. G. Tozer, M. Symons, F. McCormick, J. A. Hassel, The activity of Ets transcription factor PEA3 is regulated by two distinct MAPK cascades. Oncogene 13, 1996, 1323-1333
11. A. S. Zervos, L. Faccio, J. P. Gatto, J. M. Kyriakis, R. Brent. Mxi2, a mitogen-activated protein kinase that recognizes and phosphorylates Max protein. Proc Natl Acad Sci USA 92, 1995, 10531-10534
12. X. Z. Wang, D. Ron, Stress-induced phosphorylation and activation of the transcription factor CHOP (GADD153) by p38 MAP kinase. Science 272, 1995, 1347-1349
13. C. Fromm, O.A. Coso, S. Montaner, N. Xu, J.S. Gutkind, The small GTP-binding protein Rho links G protein-coupled receptors and Galpha 12 to the serum response element and to cellular transformation. Proc Natl Acad Sci USA 94, 1997, 10098-103
14. B.C. Kim, J.H. Kim, Exogenous C2-ceramide activates c-fos serum response element via Rac-dependent signaling pathway. Biochem J. 330,1998, 1009-14
15. R. Perona, S. Montaner, L. Saniger, I. Sanchez-Perez, R. Bravo, J. C. Lacal, Activation of Nuclear Factor-kB by Rho, CDC42, and Rac-1 proteins. Genes &Dev 11, 1997, 463-475
16. A. Sotiropoulos, D. Gineitis, J. Copeland, R. Treisman, Signal-regulated activation of serum response factor is mediated by changes in actin dynamics. Cell 98, 1999, 159-69
17. L. M. Robertson, T. K. Kerppola, M. Vendrell, D. Luk, R. J. Sineyne, C. Bocchiaro, J. Morgan, T, Curran, Regulation of c-fos expression in transgenic mice requires multiple interdependent transcription control elements. Neuron 14, 1995, 241-50
18. H. B. Sadowski, K. Shuai, J.E. Darnell Jr, M. Z. Gilmam, A common nuclear signal transduction pathway activayed by growth and cytokine receptors. Science 261, 1993, 1739-44
19. A. R. Simon, H. G. Vikis, S. Stewart, B. L. Fanburg , B. H. Cochran, K. L. Guan, Regulation of STAT3 by direct binding to the rac1 GTPase. Science 290, 2000, 144-7
20. R.P. Ballestero , P. Esteve, R. Perona , B. Jiménez , J.C Lacal (1991) Biological function of Aplysia californica rho gene. En The Superfamily of ras related Genes, pp. 237-242. NATO Advanced Science Institute Series, vol. A220. Plenum Press.
21. R. Perona , P. Esteve, B. Jimenez, R. P. Ballestero , S. Ramon y Cajal, J. C. Lacal , Tumorigenic activity of rho genes from Aplysia californica. Oncogene 8, 1993, 1285-92
22. J. L. Boerner, A. J. Danielsen, M. J. McManus, N. J. Maihle, Activation of Rho is Required For Ligand-independent Oncogenic signaling by a mutant EGF Receptor. J Biol Chem 10, 2000
23. U. G. Knaus, Rho GTPase signaling in inflammation and transformation. Immunol Res;21, 2000, 103-9
24. K. L. van Golen, Z. F. Wu, X. T. Qiao, L. W. Bao, S. D. Merajver, RhoC GTPase, a novel transforming oncogene for human mammary epithelial cells that partially recapitulates the inflammatory breast cancer phenotype. Cancer Res 60, 2000, 5832-8
25. E. A. Clark, T. R. Golub, E. S. Lander, R. O. Hynes, Genomic analysis of metastasis reveals an essential role for RhoC. Nature 406, 2000, 532-5
26. H. Suwa , G. Ohshio, T. Imamura, G. Watanabe, S. Arii, M. Imamura, S. Narumiya, H. Hiai, M. Fukumoto, Overexpression of the rhoC gene correlates with progression of ductal adenocarcinoma of the pancreas. Br J Cancer 77, 1998, 147-52
27. P. Jordan, R. Brazao, M. G. Boavida, C. Gespach, E. Chastre, Cloning of a novel human Rac1b splice variant with increased expression in colorectal tumors. Oncogene 18; 1999, 6835-9
28. G. Fritz, I. Just, B. Kaina, Rho GTPases are over-expressed in human tumors. Int J Cancer 81; 1999, 682-7
29. R. Hernandez-Alcoceba, L. del Peso, J. C. Lacal, The Ras family of GTPases in cancer cell invasion. Cell. Mol Life Sci 57, 2000, 65-76
30. L. del Peso, R. Hernandez-Alcoceba, N. Embade, A. Carnero, P. Esteve, C. Paje, JC. Lacal, Rho proteins iduce metastatic properties in vivo. Oncogene 15, 1997, 3047-3057
31. MSA. Nur-E-Kamal, J. M. Kamal, M. M. Qureshi, H. Maruta, The CDC42-specific inhibitor derived from ACK-1 blocks v-Ha-Ras-induced transformation. Oncogene 18, 1999, 7787-7793
32. R. G. Qiu, J. Chen, F. McCormick, M. Symons, A role for Rho in Ras transformation. Proc Natl Acad Sci U S A 92, 1995, 11781-5
33. R. G. Qiu, J. Chen, D. Kirn, F. McCormick, M. Symons, An essential role for Rac in Ras transformation. Nature
37, 1995, 457-9
34. S-i. Osada, M. Izawa, T. Koyama, S-i. Hirai, S. Ohno, A domain containing the Cdc42/Rac interactive binding (CRIB) region of p65PAK inhibits trnascriptional activation and cell transformation mediated by the Ras-Rac pathway. FEBS lett 404, 1997, 227-233
35. J.P. Mira, V. Bernard, J. Groffen, LC. Sanders, UG. Knaus, Endogenous, hyperactive Rac3 controls proliferation of breast cancer cells by a p21-activated kinase-dependent pathway. Proc Natl Acad Sci USA 97, 2000, 185-189
36. Y. Tang, Z. Chen, D. Ambrose, J. Liu, J. B. Gibbs, J. Chernoff, J. Field, Kinase-defective Pak1 mutants inhibit ras transformation of Rat-1 fibroblasts. Mol Cell Biol 17, 2000, 4454-4464
37. J. Westwick, Q. Lambert, G. Clark, M. Symons, L. Van Aelst, R. Pestell, C. Der, Rac regulation of transformation, gene expression, and actin reorganization by multiple, PAK-independent pathways. Mol Cell Biol 17, 1997, 1324-1335
38. Y. Tang, S. Marwaha, J. L. Rutkowski, G. I. Tenekoon, P. C. Philips, J. Field. A role for Pak protein kinases in Schwann cell transformation, Proc Natl Acad Sci USA 95, 1998, 5138-5144
39. H. Teramoto, O. Coso, H. Miyata, T. Igishi, T. Miki, S. Gutkind, Signaling form the small GTP-binding proteins Rac1 and Cdc42 to the c-Jun N-terminal kinase/Stress-activated protein kinase pathway. J Biol Chem 271, 1996, 27225-27228
40. J. Brown, L. Stowers, M. Baer, J. Trejo, S. Coughlin, J. Chant, Human Ste20 homolog hPAK1 links GTPases to the JNK MAP kinase pathway. Curr. Biol 6, 1996, 598-605
41. S. Bagrodia, R. Derijard, R. Davis, R. Cerione, Cdc42 and PAK-mediated signlaing leads to Jun Kinase and p38 mitogen-activated protein kinae activation, J Biol Chem 270, 1995, 27995-27998
42. S. Zhang, J. Han, M. Sells, U. Chernoff, U. Knaus, R. Ulevitch, G. Bokoch, Rhofamily GTPases regulates p38 mitogen-activated protein kinase through the downstream mediator Pak1. J Biol Chem 270, 1995, 23934-23936
43. N. Tapon, K-i. Nagata, N. Lamarche, A. Hall, A new Rac target POSH is an SH3-containing scaffold protein involved in the JNK and NF-kB signalling pathways. EMBO J 17, 1998, 1395-1404
44. H. Teramoto, O. Coso, H. Miyata, T. Igishi, T. Miki, S. Gutkind, Signaling form the small GTP-binding proteins Rac1 and Cdc42 to the c-Jun N-terminal kinase/Stress-activated protein kinase pathway. J Biol Chem 271,1996, 27225-27228
45. H. Teramoto, P. Crespo, O.A.Coso, T. Igishi, N. Xu, J.S. Gutkind, The small GTP-binding protein rho activates c-jun N-terminal kinases/stress-activated protein kinases in human kidney 293T cells. Evidence for a PAK-independent signaling pathway. J Biol Chem 271, 1996, 25731-25734
46. P. Gerwins, J.L. Blank, G.L. Johnson, Cloning of a novel mitogen-activated protein kinase kinase kinase, MEKK4m that selectively regulates the c-jun amino terminal kinase pathway. J Biol Chem 272, 1997, 8288-8295
47. L. Tibbles, Y. Ling, F. Kiefer, J. Chan, N. Iscove, J.R. Woodget, N.J. Lassam, MLK-3 activates JNK/SAPK and p38/RK pathways via SEK1 and MKK3/6. EMBO J 15, 1996, 7026-7035
48. T. Leung, E. Manser, L. Tan, L. Lim, A novel Serine/threonine kinase binding the Ras-related RhoA GTPases which translocates the kinase to peripheral membranes. J Biol Chem 270, 1995, 29051-29054
49. S. P. Henner, T. G. Hofman, A. Ushmorov, O. Dienz, I. Wing-Lan Leung, N. Lassam, C. Sheidereit, W. Droge, M. L. Schmitz, Mixed-lineage kinase 3 delivers CD3/CD28-derived signals into the IkB -kinase complex. Mol Cell Biol 20, 2000, 2556-2568
50. S. Huang, L. New, Z. Pan, J. Han, G. R. Nemerow. Urokinase plasminogen activator/urokinase-specific surface receptor expression and matrix invasion by breast cancer cells requires constitutive p38alpha mitogen-activated protein kinase activity. J Biol Chem 275, 2000, 12266-12272
51. L. Montero, Y. Nagamine, Regulation by p38 mitogen-activated protein kinase of adenylate- and urydilate-rich element-mediated urokinase-type plasminogen activator (uPAR) messenger RNA stability and uPA-dependent in vitro cell invasion. Cancer Res 59, 1999, 5286-5293
52. S. M. Muller, E. Okan, P. Jones, Regulation of urokinase receptor transcription by Ras- and Rho-family GTPases. Biochem Biophys Res Commun 270, 2000, 892-898
53. S. Bourdoulous, G. Orend, D. A. MacKenna, R. Pasqualini, E. Ruoslahti. Fibronectin matrix regulates activation of RHO and CDC42 GTPases and cell cycle progression. J Cell Biol 143, 1998, 267-276
54. J. Dang, D. Boyd, H. Wang, H. Allgayer, W. F. Doe, Y. Wang, A region between -141 and -61 containing a proximal AP-1 is essential for contitutive expression of urokinase-type plasminogen activator receptor. Eur J Biochem 264, 1999, 92-99
55. Y. Wang, J. dang, H. Wang, H, Allgayer, G. A. Murrell, D. Boyd, Identification of a novel factor-kappa B sequence involved in expression of urokinase-type plasminogen activator receptor. Eur J Biochem 267, 2000, 3248-3254
56. D. J. Sulciner, K. Irani, Z. X. Yu, V. J. Ferrans, C. P. Goldschmidt, T. Finkel, Rac1 regulates a cytokine-stimulated redox-dependent pathway necessary for NF-kB activation. Mol Cell Biol 16, 1996, 7115-7121
57. T.S. Finco, J. K. Westwick, J. L. Norris, A. A. Beg, C. J. Der, A. S. Baldwin Jr, Oncogenic Ha-Ras-induced signaling activates NF-kappaB transcriptional activity, which is required for cellular transformation. J Biol Chem 272, 1997, 24113-24116
58. M. Arsuna, F. Mercurio, A. L. Oliver, S. S. Thorgeirsson, G. E. Sonenshein. Role of the IkappaB kinase complex in oncogenic Ras- and Raf-mediated transformation of rat liver epithelial cells. Mol Cell Biol 20, 2000, 5381-5391
59. B. Baumann, C. K. Weber, J. Troppmai, S. Whiteside, A. Israel, U.R. Rapp, T. Wiryh. Raf induces NF-kappaB by membrane shuttle kinase MEKK1, a signaling pathwya critical for transformation. Proc Natl Acad Sci USA97, 2000, 4615-4620
60. H. Jo, R. Zhang, H. Zhang, T. A. McKinsey, J. Shao, R. D. Beauchamp, C. W. Ballard, P. Liang, NF-kappa B is required for H-ras oncogene induced abnormal cell proliferation and tumorigenesis. Oncogene 19, 2000, 841-849
61. J. Y. Reuther, G. W. Reuther, D. Cortez, A. M. Pendergast, A. S. Baldwin Jr, A requirement for NF-kappaB activation in Bcr-Abl-mediated transformation. Genes &Dev 12, 1998, 968-981
62. I. P. Whitehead, Q. T. Lambert, J. A. Glaven, K. Abe, K. L. Rossman, G. M. Mahon, J. M. Trzaskos, R. Kay, S. L. Campbell, C. J. Der. Dependence of Dbl and Dbs transformation on MEK and NF-kB activation. Mol Cell Biol 19, 1999, 7759-70
63. P. Frankel, M. Ramos, J. Flom, S. Bychenok, T. Joseph, E. Kerkhoff, U. R. Rapp, L. A. Feig, D. A. Foster, Ral and Rho-dependent activation of phospholipase D in v-Raf-transformed cells. Biochem Biophys Res Commun 255, 1999, 502-507
64. E. Sahai, T. Ishizaki, S. Narumiya, R. Treisman, Transformation mediated by Rho requires activity of ROCK kinases. Curr biol 9, 1999, 136-145
65. T. Ishizaki, M. Uehata, I. Tamechika, J. Keel, K. Nonomura, M. Maekawa, S. Narumiya, Pharmacological properties of Y-27632, a specific inhibitor of Rho-Associated Kinase. Mol Phar 57, 2000, 976-983
66. K. Itoh, K. Yoshioka, H. Akedo, M. Uehata, T. Ishizaki, S. Narumiya, An essential part for Rho-associated kinase in the transcellular invasion of tumor cells. Nat Med 5, 1999, 221-225
67. T. Goi, M. Shipitsin, Z. Lu, D. A. Foster, S. G. Klinz, L. A. Feig, An EGF receptor/Ral-GTPase signaling cascade regulates c-Src activity and substrate specificity. EMBO J 19, 2000, 623-30
68. C. S. Hill, J. Wynne, R. Treisman, The Rho family GTPases RhoA, Rac1, and CDC42Hs regulate transcriptional activation by SRF. Cell 81, 1995, 1159-1170
69. S. Montaner, R. Perona, L. Saniger, J. C. Lacal, Activation of serum response factor by RhoA is mediated by the nuclear factor-kB and C/EBP transcription factors. J Biol Chem 274, 1999, 8506-8515
70. A. S. Alberts, O. Geneste, R. Treisman, Activation of SRF-regulated chormosomal templates by Rho-family GTPases requires a signal that also induces H4 hyperacetylation. Cell 92, 1998, 475-487
71. P. F. Lebowitz, W. Du, G. C. Prendergast, Prenylation of RhoB is required for its cell transforming function bu not its ability to activate serum response element-dependent transcription. J Biol Chem 272, 1997, 16093-16095
72. E. Sahai, A. S. Alberts, R. Treisman, RhoA effector reveal distinct effector pathways for cytoskeletal reorganization, SRF activation and transformation. EMBO J 17, 1998, 1350-1361
73. M. Zohar, H. Teramoto, B. Z. Katz, K. M. Yamada, J. S. Gutkind, Effector domain mutants of Rho dissociated cytoskeletal changes from nuclear signaling and cellular transformation. Oncogene 17, 1998, 991-998
74. B. Jiménez, M. Arends, P. Esteve, R. Perona, Y. Sanchez, S. Ramón y Cajal, A. Wyllie, and J.C. Lacal, Induction of apoptosis in NIH3T3 cells after serum deprivation by overexpression of rho-p21, a GTPase protein of the ras superfamily. Oncogene 10, 1995, 811-816
75. Y. Y. Kong, K. D. Fischer, M. F. Bachmann, S. Mariathasan, I. Kozieradzki, M. P. Nghiem, D. Bouchard, A. Bernstein, P. S. Ohashi, J. M. Penninger, Vav regulates peptide-specific apoptosis in thymocytes. J Exp Med 188, 1998, 2099-111
76. P. Lores, L. Morin, R. Luna, and G.Gacon, Enhanced apoptosis in the thymus of transgenic mice expressing constitutively activated formas of human Rac2GTPase. Oncogene 15, 1997, 601-605
77. P. Esteve, N. Embade, R. Perona, B. Jiménez, L. del Peso, J. León, M. Arends, T. Miki, and J.C. Lacal, Rho-regulated signals induce apoptosis in vitro and in vivo by a p53-independent, but Bcl-2 dependent pathway. Oncogene 17, 1998, 1855-1869
78. N. Embade, P. F. Valerón, S. Aznar, E. López-Collazo, J. C. Lacal, Apoptosis induced by rac GTPase correlates with induction of FasL and ceramides production. Mol Biol Cell 11, 2000, 4347-58
79. T. Rudel, G. M. Bokoch, Membrane and morphological changes in apoptotic cells regulated by caspase-mediated activation of PAK2. Science 276, 1996, 1571-1574
80. N. Lee, H. MacDonald, C. Reinhardt, R. Halenbeck, A. Roulston, T. Shi, L. T. Williams, Activation of hPAK65 by caspase cleavage induces some of the morphological changes of apoptosis. Proc Natl Acad Sci USA 94, 1997, 13642-13647
81. M. Takahashi, H. Mukai, M. Toshimori, M. Miyamoto, Y. Ono, Proteolytic activation of PKN by caspase-3 related protease during apoptosis. Proc Natl Acad Sci USA 95, 1998, 11566-11571
82. H. Koh, K. H. Lee, D. Kim, S. Kim, J. W. Kim, J. Chung, Inhibition of Akt and its anti-apoptic activities by tumor necrosis factor-induced PRK2 cleavage. J Biol Chem 275, 2000, 34451-8
83. T. H. Chuang, K. M. Hahn, J. D. Lee, D. E. Danley, G.M. Bokoch, The small GTPase Cdc42 initiates an apoptotic signaling pathway in Jurkat T lymphocytes. Mol Biol Cell. 9, 1997, 1687-98
84. E. Gulbins, K.M. Coggeshall, B. Brenner, K. Schlottmann, O. Linderkamp, and F. Lang, Fas-induced apoptosis is mediated by activation of a Ras and Rac protein-regulated signalling pathway. J. Biol. Chem. 271, 1996, 26389-26394

Figure 1
Signal transduction pathways involved in RhoA-, Rac1- and Cdc42-mediated regulation of transcription.

Figure 2
TFs and effectors to Rho GTPases known to be involved in transformation/metastasis and programmed cell death.

Back . . .